Pathogenicity of Ganoderma boninense tested by inoculation of oil palm seedlings

The pathogenicity of Ganoderma boninense isolated from oil palm was tested using wood blocks from a rubber tree as the substrate for the inoculum. Seedlings with roots placed in close contact with the inoculum blocks gave 100% infection. There was a progressive desiccation of leaves from the oldest to the youngest. Sporophores emerged as a white mass of mycelial tissue which rapidly became button-like. This was followed by the development of the stipe. Bracket-shaped sporophores formed within 12–16 weeks after inoculation. Longitudinal sections of affected seedlings revealed bole and stem decay. Hyphae were detected in the cortex, endodermis, pericycle and vascular tissues of infected roots. Gummosis and the production of tyloses in xylem vessels were common.     

Effect of soil solarization on the control of Phytophthora root rot in avocado

Phytophthora root rot is of paramount importance in avocado orchards of southern Spain. Soil solarization has been demonstrated to control the pathogen in infested areas from which infected trees had been removed. We aimed to determine whether soil solarization in established avocado orchards controls the disease. Soil solarization increased average maximum hourly soil temperatures by 6.5–6.9°C in unshaded areas of avocado orchards in coastal areas of southern Spain, depending on depth and year. The corresponding temperatures in shaded areas were c. 2–3°C lower. P. cinnamomi in soil, on infected avocado rootlets, and in a nutrient substrate buried at 30–60 cm depth was reduced to negligible amounts after 6–8 weeks of solarization in both unshaded and shaded locations of avocado orchards. P. cinnamomi could not be detected in avocado rootlets up to 14 months later, suggesting a long-term effect. Soil solarization did not affect growth of the trees, and fruit yields were increased as compared with control plots. Following soil solarization for 3 weeks from mid-July 1994, when maximum hourly temperatures reached 33–36°C, P. cinnamomi could not be recovered from a depth of up to 45 cm in unshaded areas or from a depth of up to 30 cm in shaded areas after the initial 10-day period. The viability of inoculum of the pathogen buried at depths between 15 and 60 cm in bare soil was determined by sequential sampling in two solarization experiments starting 12 June and 4 July 1995, respectively. In the first experiment, P. cinnamomi could not be detected at any depth after 4–8 weeks of solarization in unshaded areas but could be recovered at all depths except 15 cm in shaded areas. In the second experiment, where temperatures were higher and the soil surface not shaded, P. cinnamomi could not be recovered after 2 weeks at 15 and 30 cm.     

Factors affecting the incidence and severity of Spongospora subterranea infection and galling in potato roots

The incidence and severity of root infection and root galling caused by Spongospora subterranea were assessed in potato plants (cv. Estima) grown under controlled environmental conditions. The effects of temperature, soil type, soil moisture regime and soil inoculum level on infection and root gall development were determined by molecular and visual methods at two plant growth stages. Root gall severity was scored at harvest, after which DNA was extracted from the roots and quantified in a real-time polymerase chain reaction (PCR) assay specific for S. subterranea . Root galling was severe at 17°C, with a disease score of 3·1 on a 0–4 scale, low (0·6) at 12°C, and did not occur at 9°C. The level of inoculum in soil, in the form of artificially added sporosori, had no effect on the incidence and severity of visual symptoms, with 21%, 41% and 33% incidence observed at 5, 15 and 50 sporosori g⁻¹ soil, respectively. Incidence of infection, as detected by the real-time PCR assay, was greater with increasing soil inoculum concentrations, ranging from 48% at 5 sporosori g⁻¹ to 59% (15 sporosori g⁻¹) and 73% (50 sporosori g⁻¹) of plants infected at maturity, but this effect was not statistically significant. No correlation was found between the occurrence of galls on roots and powdery scab on tubers of the same plants.     

Eradication of Plasmodiophora brassicae during composting of wastes

Survival of infectious inoculum of the clubroot pathogen Plasmodiophora brassicae was assessed following bench-scale flask composting experiments and large-scale composting procedures. Clubroot-affected material was provided by artificial inoculation of Chinese cabbage or naturally infected Brussels sprout and cabbage roots. Both sets of diseased material were used in flask experiments, and the latter in large-scale windrow and aerated tunnel experiments. Municipal green wastes, onion waste and spent mushroom compost were evaluated in flask experiments with varying temperature, aeration and moisture conditions. Green wastes were used in larger-scale composts. Within the limits of a Chinese cabbage seedling bioassay, both temperature and moisture content were critical for eradication of P. brassicae spores extracted from composted clubroot-affected residues. Incubation in compost at 50°C for 7 days or 1 day at 60°C with high moisture levels (= −5 kPa matric potential or 60% w/w moisture content) eradicated inoculum from artificially inoculated Chinese cabbage roots. In large-scale windrows and aerated tunnels, the pathogen was eradicated from naturally infected brassica wastes after 6–7 days at 54–73°C.     

UNTERSUCHUNGEN ZUR BIOLOGIE UND ÖKOLOGIE DER HÜHNER-HIRSE ECHWOCHLOA CRUS-GALLI L. BEAUV.

Summary. 1. Echinochloa crus-galli is a typical grass weed of root crops in warmer regions, germinating late and thus covering the soil at a late stage. It originates in central and east Asia and is now a weed of world-wide importance.
2. High temperatures are necessary for germination which begins in spring, but not before the soil temperature reaches 15° C. Minimum, optimum and maximum germination temperatures of 13° C, 20–30° C and 40° C were found in S.W. Germany.
3. Seeds of E. crus-galli are dormant during the first 3–4 months after harvest. Those developing relatively early in the growing season require rather longer for after-ripening than seeds which mature later.
4. For optimum germination, water saturation of the soil of 70–90% is required.
5. Soil acidity has some influence, and there is an apparent germination optimum around neutrality. Light also induces germination.
6. Seeds can emerge from a relatively wide range of depths. Greatest emergence and the strongest plants resulted from seeds at 2–6 cm, but even from 10 cm a high percentage of seedlings is likely to emerge.
7. Further development proceeds rapidly. The first panicles arc already formed 6–7 weeks after emergence in favourable conditions, but full maturity is possible only if there are high temperatures in late summer.
8. E. crus-galli is a hygrophilous species, with best development on medium heavy soil, sandy loam or loamy sand with sufficient water supply.
9. E. crus-galli is indifferent to the lime content of the soil.
10. Best development occurs with high fertility, and a rich supply of nitrogen is especially important.
Recherches sur la biologie et écologie du panic Echinochloa crus-galli (L.) Beauv.     

Genetic structure of a population of Ganoderma boninense on oil palm

Ganoderma boninense (the causal agent of basal stem rot of oil palm in Papua New Guinea) has a tetrapolar mating system with multiple alleles. Investigations into the population structure of G. boninense , using interfertility between isolates as a marker, revealed that the population on oil palm was comprised predominantly of genetically distinct individuals, although a number of isolates were found to share single mating alleles. No direct hereditary relationship was found between isolates on neighbouring or spatially separated diseased palms, indicating that outcrossing had probably occurred over several generations in the founder population prior to colonization of oil palm. In this study, a total of 81 A and 83 B mating type alleles (factors) were detected with 18 allelic repeats at the A locus and 17 at the B locus. Alleles appeared to be randomly dispersed throughout the population in each study block, although there was a significantly ( P  < 0·01) higher proportion of allelic repeats between estates separated by several kilometres compared to within estates. This is the first study on the population structure of this species and confirms that sexual reproduction is a significant component of the epidemiology of this disease.     

The influence of inoculum concentration, relative humidity, and temperature on infection of greenhouse tomatoes by Botrytis cinerea

Botrytis cinerea causes serious crop losses in greenhouse tomato crops through infection of flowers and stem wounds. Experiments were carried out to determine the effects of inoculum concentration, relative humidity (RH), and temperature at these two infection sites. Infection of permanent flower parts increased as a function of inoculum concentration and both length of exposure to high RH (approximately 100% for 0–36 h) and specified continuous RH (56–100%). A low level of infection was still evident under continuous 56% RH. Interruption of periods of high RH with breaks of low RH did not reduce infection. Infection of stem wounds was less dependent on inoculum concentration or RH. Factorial combinations of inoculum concentration, RH, and temperature produced significant interactions. Higher temperature increased infection of flowers but reduced infection of stem wounds. The main implications for control in commercial crops are as follows. Lowering the aerial spore concentration by maintaining the disease at a low level will reduce flower infection. Lowering RH will reduce but not eliminate flower infection but will have only a small effect on stem infection. Raising the temperature (from 15 to 25°C) will reduce stem infection, and whilst flower infection increases, this is counteracted by increased flower production and a decrease in the proportion of infections reaching the peduncle and stem.     

Infection of sugar beet by Polymyxa betae in relation to soil temperature

The effects of soil temperature on infection of sugar-beet roots by the soil-borne fungus Polymyxa betae were investigated in controlled environments. Pre-germinated seeds were sown in pots of naturally infested soil and seedlings sampled at frequent intervals over a period of several weeks. Within the range 10-30°C, the optimum soil temperature for infection was c. 25°C; the time between sowing and the first detectable infection was shortest and the subsequent rate of infection most rapid at this temperature. No infection was observed over 80 days at 10°C.
Both root and shoot dry weight were reduced on plants growing in infested soil at 15, 20 and 25 C compared with those growing in uninfested soil. In general, root growth was more severely affected than shoot growth and the effects were most pronounced at 20°C. These results were confirmed in a subsequent experiment in which P. betae -infected root material was used as the inoculum. In addition to its role as the vector of beet necrotic yellow vein virus (the cause of Rhizomania disease), the significance of P. betae as a plant pathogen in its own right is discussed.     

Detection of Colletotrichum coccodes and Helminthosporium solani in soils by bioassay

The sensitivity of a bioassay in detecting soil inoculum of Colletotrichum coccodes and Helminthosporium solani was examined using potato minitubers and microplants. Tests were conducted on soils which were collected from fields in which the interval after a previous potato crop differed, and which were also artificially infested with conidia or microsclerotia. For C. coccodes , determining plant infection based on the occurrence of infected roots after 9–12 weeks was a sensitive method for detecting and quantifying the amount of inoculum in soil. Infestations of less than 0·4 microsclerotia per g soil were detected in artificially infested soils. A semiselective medium, developed for isolating C. gloeosporioides from pepper, detected soil infestations by C. coccodes as low as nine conidia or one microsclerotium per g soil in artificially infested soil. For H. solani , infection on minitubers was a sensitive measure, with soil inoculum of fewer than 10 conidia per g soil being detected. Soil infestation could be quantified by assessing the percentage surface area of minitubers covered by sporulating lesions, which was strongly related to the amount of soil infestation. The results of these bioassay tests were compared with published results for real-time quantitative PCR assays on the same soils. The two methods were in good agreement in artificially infested soils, but the bioassay appeared to be more sensitive with naturally infested soils.     

Biological control of damping-off of sugar beet by Pseudomonas putida applied to seed pellets

Pseudomonas putida 40RNF applied to seed pellets reduced the occurrence of Pythium damping-off of sugar beet. A density of 6 × 10⁷ 40RNF per pellet reduced Pythium damping-off from 70 to 26% when seeds were sown in artificially infested soil (250 propagules Pythium ultimum per g dry soil). The efficacy of 40RNF was dependent on its density in the seed pellet (in the range 2 × 10⁴–6 × 10⁸ per pellet) and on the number of propagules of Pythium in soil. 40RNF declined to or stabilized at approximately 1 × 10⁶ per pellet 3 days after planting, and this was independent of the inoculum density. This indicated that the crucial steps resulting in damping-off of sugar beet caused by Pythium ultimum must occur within 3–4 days of sowing. 40RNF reduced pericarp colonization by P. ultimum by 43% 48 h after planting and caused a 68% decrease in the number of sporangia of P. ultimum in the surrounding soil (0.0–5.0 mm). P. putida 40RNF also reduced pre and post-emergence damping-off (from 69.5 to 37.5%) caused by indigenous populations of Pythium species in an infested soil and this was as effective as the fungicide hymexazol (69.5 to 40%).     

Basal stem rot of oil palm (Elaeis guineensis); mode of root infection and lower stem invasion by Ganoderma boninense

Reproducible infection of intact roots of oil palm ( Elaeis guineensis ) with Ganoderma boninense , the cause of basal stem rot, showed penetration followed by rapid longitudinal progression of hyphae and colonization of the lower stem (bole). Light and transmission electron microscopy showed invasion of the root cortex, with no evidence of selective progression through the vascular system or lacunae. In newly colonized tissue the fungus behaved as a hemibiotroph, with numerous, wide, intracellular hyphae occupying entire host cells that possessed intact cell walls and contained discernible cytoplasm and organelles. In the bole this phase coincided with a complete depletion of previously abundant starch grains in advance of invasion. Subsequently, in the roots and colonized stem base, widespread necrotrophic, enzymatic attack of all layers of the host cell walls occurred. Hyphae were intra- and intercellular and intramural and associated host cell wall degradation was often at a distance from hyphae, resulting in cavities within cell walls. A third developmental stage was the formation of an extensive, melanized, tough mycelium or pseudo-sclerotium which surrounded roots and comprised many very thick-walled cells encasing more typical thin-walled hyphae. Macroscopic observation of and isolation from the bole of randomly felled, commercial palms provided confirmatory evidence that multiple infections originated in the roots before spreading into the base of long-established palms.     

Effects of temperature and moisture on disease and fruit body development of Mycosphaerella pinodes on pea (Pisum sativum)

The effects of temperature (5–30°C) and the duration of moisture on the development of ascochyta blight ( Mycosphaerella pinodes ) on pea seedlings, grown under controlled conditions, were investigated. The optimum temperature for monocyclic processes was 20°C. At this temperature, pycnidiospores germinated after 2 h, appressoria formed after 6 h and the germ-tube penetrated the leaf cuticle after 8 h. Disease symptoms were evident after 1 day of incubation and the first pycnidia formed after 3 days. Longer wetting periods were required for disease development and pycnidial formation at non-optimal temperatures. Disease severity and the number of pycnidia formed on leaves increased with temperature from 5 to 20°C, then decreased between 20 and 30°C. Polynomial equations were fitted to predict the stages of infection, incubation, latency and disease development as functions of temperature and duration of moisture. These equations allow comparisons of pathogen spread with plant development and could be incorporated into disease development models used for crop management programmes.     

Solarization of soil in piles for the control of Meloidogyne incognita in olive nurseries in southern Spain

The potential of solarization to control Meloidogyne incognita in piles of soil used at olive nurseries in southern Spain was studied in 1999 and 2000. Kaolin and soil infested with free eggs and egg masses of the nematode in nylon bags were buried 20 and 40 cm deep inside conical piles of soil 80 cm high and with a base diameter of 1 m. Soil piles were solarized for 3 weeks in July and August. The effect of various periods of solarization was assessed by egg hatch bioassays in sterile water, and by infectivity to tomato plants. Maximum soil temperature at 20 cm depth in solarized piles was 47·4°C in 1999 and 48·2°C in 2000, compared with 32·9°C and 31·7°C in nonsolarized piles. Solarization reduced egg hatch by > 95% compared with nonsolarized samples, irrespective of type, burial depth and location of inocula in a soil pile. Egg hatch of egg mass-infested samples buried at 20 cm depth was higher than that of free eggs buried at the same depth. The differential effect associated with burial depth and type of inoculum was not found in solarized piles. In nonsolarized piles, hatch of free eggs from samples buried at 40 cm depth was higher than that from samples buried at 20 cm depth. Egg hatch in samples from solarized piles was lower than that from nonsolarized piles. A bioassay of tomato plants in 2000 confirmed the reduction in infectivity of free eggs buried in solarized soil piles. Under the conditions in southern Spain, solarization of 40 cm-high piles of soil for 3 weeks can therefore be used for the control of root-knot nematodes in potting soil for olive nursery production.     

In vitro effects of temperature and wet periods on infection of oilseed rape by Alternaria brassicae

The influence of temperature and duration of wet periods on infection of oilseed rape by Alternaria brassicae was studied on detached leaves and pods, leaf disks and intact seedlings. Infections increased with age of leaf and the interaction between temperature and leaf age was highly significant. On older leaves infection was optimal at 25°C. There were many infections also at 15, 20 and 29 C but relatively few infections at 10°C. On pods most infections were observed at 20 C, the highest temperature studied. Infection at each temperature increased progressively with duration of surface wetness. The minimum wet periods for infection of leaves were 3 h at 20–25°C, 4 h at 15°C, 6–9 h at 10 C and 12–24 h at 5 C and for infection of pods, between 6 h and 9 h at 10°C and 6 h (or less) at 15°C and 20 C. On leaves, dry periods interrupting wet periods limited lesion development to that obtained with the initial wet period only; on pods some further infections developed when pods were re-wetted. Dry periods of 3 h and 6 h following the inoculation of pods reduced subsequent infection but there was no further reduction by longer periods of drying to 48 h.     

Quantitative relationships between inoculum of Melampsora larici-epitea and corresponding disease on Salix

Six Salix clones were inoculated with urediniospores of four isolates of Melampsora larici-epitea at five inoculum levels using a leaf-disc method. Disease reactions were recorded using a digital camera; the number and size of uredinia were examined using image analysis software; and spore yield per leaf disc was measured. In three Salix / Melampsora combinations, S.  ×  mollissima 'Q83'/Q1 (LET4); S. viminalis '78183'/V1 (LET1); and S.  ×  stipularis /V1, pustule numbers increased as inoculum density became higher. In the remainder, S. viminalis 'Mullatin'/V1; S.  ×  calodendron /DB (LET3); and S. burjatica 'Korso'/K (LR1), pustule numbers initially increased, then decreased as inoculum densities exceeded 140–360 spores per disc. Calculated infection efficiency ranged from 0·11 to 0·20 on the three willows inoculated with V1: 0·16–0·68 for S.  ×  calodendron /DB; 0·20–0·55 for 'Q83'/Q1; and 0·07–0·48 for Korso/K. In single-spore inoculations, up to 10% of spores produced single uredinia. Infection efficiency increased sharply between inoculum densities of 1–40 spores per leaf disc. Spore yield was more closely correlated to pustule area (accounting for 61·2% variance for the combined data) than to the number of pustules (42·7% variance). For spore yields in relation to pustule numbers, clone-specific individual lines having different intercepts and slopes fitted significantly better than either a single line for all the tested willows, or parallel lines fitted to each clone ( P  < 0·001). For spore yields in relation to pustule area, clone-specific individual parallel lines were significantly better than a single line ( P  < 0·001).     

氟乐灵处理后棉苗对枯萎病菌的抗侵入及抗扩展特性

 以1μg/g土的氟乐灵播前土壤处理,棉花出苗后移栽到无氟乐灵的土壤中并接种棉花枯萎病菌(Fusarium axysporum f.sp.vasinfectum)。结果表明氟乐灵处理明显降低了棉苗枯萎病的株发病率和病情指数,提高了棉苗对枯萎病的抗病性。氟乐灵处理组棉苗根部和茎部枯萎病菌侵染率均低于对照组,特别是上部茎片段中处理组侵染率降低更为明显。氟乐灵处理组棉苗根部和茎部的菌量明显降低,而且处理组茎部菌量的增长明显滞后于对照。以上结果表明,病菌对氟乐灵处理棉苗根部的侵入和病菌在棉苗体内的纵向扩展及增殖受到了阻抑,因而氟乐灵处理后棉苗对枯萎病菌的抗性表现了抗侵入和抗扩展的特性。试验还证明,氟乐灵处理并接种病菌的棉苗茎部组织中类萜烯醛的含量明显提高;并产生一种能抑制病菌大型分生孢子萌发的真菌毒性物质。这些抑菌物质的产生和积累可能与抗扩展特性有关。     

氟乐灵处理后棉苗对枯萎病菌的抗侵入及抗扩展特性

以1μg/g土的氟乐灵播前土壤处理,棉花出苗后移栽到无氟乐灵的土壤中并接种棉花枯萎病菌(Fusarium axysporum f. sp. vasinfectum)。结果表明氟乐灵处理明显降低了棉苗枯萎病的株发病率和病情指数,提高了棉苗对枯萎病的抗病性。氟乐灵处理组棉苗根部和茎部枯萎病菌侵染率均低于对照组,特别是上部茎片段中处理组侵染率降低更为明显。氟乐灵处理组棉苗根部和茎部的菌量明显降低,而且处理组茎部菌量的增长明显滞后于对照。以上结果表明,病菌对氟乐灵处理棉苗根部的侵入和病菌在棉苗体内的纵向扩展及增殖受到了阻抑,因而氟乐灵处理后棉苗对枯萎病菌的抗性表现了抗侵入和抗扩展的特性。试验还证明,氟乐灵处理并接种病菌的棉苗茎部组织中类萜烯醛的含量明显提高;并产生一种能抑制病菌大型分生孢子萌发的真菌毒性物质。这些抑菌物质的产生和积累可能与抗扩展特性有关。     

Effect of soil temperature on disease development in melon plants infected by Monosporascus cannonballus

The effect of soil temperature on melon collapse induced by Monosporascus cannonballus was studied in the laboratory and in the field. In the laboratory, ascospore germination and hyphal penetration into melon roots were enhanced by increasing the temperature from 20 to 32°C. The optimum temperature for mycelial growth of five isolates of M. cannonballus was 30°C. In the field, the effect of temperature was studied in experiments conducted during the winter and autumn cropping seasons from 1995 to 1998. Disease progress was much faster in the autumn than in the winter crop seasons. Disease incidence reached 100% in the three consecutive autumn seasons studied. In the winter seasons, however, planting date influenced disease incidence. Early planting, at the beginning of January, resulted in a low disease incidence (6–26%, 125 days after planting), whereas planting at the end of January resulted in higher disease incidence (72–88%, 95–119 days after planting). In plots in which the soil was artificially heated to 35°C during the winter season, disease incidence reached 85%, as in the autumn season. Plants grown during the winter in unheated soil, or in artificially heated soil disinfected with methyl bromide, did not collapse. Root colonization by the pathogen was higher in the autumn and in heated soil than in the winter season in nonheated soil. Fifty per cent of root segments were colonized 35, 42 and 67 days after planting in the winter-heated, autumn and winter-unheated plots, respectively. A high correlation was found between soil temperatures above 20°C during the first 30 days after planting and disease severity. It is suggested that soil temperature during the early stages of plant development is an important factor in disease development and the expression of melon collapse caused by M. cannonballus.     

Sporulation of Pyrenopeziza brassicae (light leaf spot) on oilseed rape (Brassica napus) leaves inoculated with ascospores or conidia at different temperatures and wetness durations

In controlled environment experiments, sporulation of Pyrenopeziza brassicae was observed on leaves of oilseed rape inoculated with ascospores or conidia at temperatures from 8 to 20°C at all leaf wetness durations from 6 to 72 h, except after 6 h leaf wetness duration at 8°C. The shortest times from inoculation to first observed sporulation ( l ₀), for both ascospore and conidial inoculum, were 11–12 days at 16°C after 48 h wetness duration. For both ascospore and conidial inoculum (48 h wetness duration), the number of conidia produced per cm² leaf area with sporulation was seven to eight times less at 20°C than at 8, 12 or 16°C. Values of Gompertz parameters c (maximum percentage leaf area with sporulation), r (maximum rate of increase in percentage leaf area with sporulation) and l ₃₇ (days from inoculation to 37% of maximum sporulation), estimated by fitting the equation to the observed data, were linearly related to values predicted by inserting temperature and wetness duration treatment values into existing equations. The observed data were fitted better by logistic equations than by Gompertz equations (which overestimated at low temperatures). For both ascospore and conidial inoculum, the latent period derived from the logistic equation (days from inoculation to 50% of maximum sporulation, l ₅₀) of P. brassicae was generally shortest at 16°C, and increased as temperature increased to 20°C or decreased to 8°C. Minimum numbers of spores needed to produce sporulation on leaves were ≈25 ascospores per leaf and ≈700 conidia per leaf, at 16°C after 48 h leaf wetness duration.     

Genetic diversity of Ganoderma in oil palm plantings

Isolates of Ganoderma spp. associated with basal stem rot of oil palm were obtained by sampling from two palm plantings in Malaysia. Members of a subset of these were compared using a combination of somatic incompatibility and molecular genetic analyses. Incompatibility interactions between heterokaryons in culture indicated that Ganoderma generally occurred as numerous distinct strains, with compatible strains usually confined to individual palms. Analysis of restriction fragment length polymorphisms (RFLPs) in presumptive mitochondrial DNA (mtDNA) revealed heterogeneity among isolates from neighbouring palms, and from within individual palms. The combination of mtDNA variability and somatic incompatibility suggests that Ganoderma BSR does not spread by direct root-to-root contact between palms. The results may indicate infection spread through basidiospore dispersal or through contact with a heterogeneous long-term residual inoculum in debris.